Anderl, Ines

Abstract [en]

Blood cells have important roles in immune reactions in all metazoan species. In Drosophila melanogaster larvae, phagocytic plasmatocytes are the main blood cell (hemocyte) type. Lamellocytes participate in encapsulating foreign objects and are formed in response to parasitoid wasps laying their eggs into the hemocoel of the larvae. The immune reaction against wasps requires controlled recruitment and action of hemocytes from the lymph glands, sessile islets and circulation. However, the contribution of these different hematopoietic compartments to the immune-induced hemocyte pool remains unclear. We used eater-GFP and MSNF9MO-mCherry to fluorescently tag plasmatocytes and lamellocytes, respectively, and utilized flow cytometry and in vivo imaging to assess the hemocyte numbers and types in circulation and in sessile compartments after infection by three wasp species of the genus Leptopilina. We detected five different hemocyte types based on fluorescence, and a population of non-fluorescent cells. While non-infected larvae generally had only one, eaterGFP-high plasmatocyte population, early after wasp infection a new, eaterGFP-low cell population appeared in circulation. EaterGFP-high and –low cells both accumulated msnCherry during the immune response, and formed two cell lineages. Whereas the eaterGFP-low cells gradually lost GFP, the eaterGFP-high cells retained it at high levels. We suggest that eaterGFP-low cells represent an immune-induced hemocyte precursor cell pool, which, via a prelamellocyte stage, gives rise to lamellocytes. EaterGFP-high plasmatocytes also differentiated into large, msnCherry-positive hemocytes on wasp eggs, but these cells retain plasmatocyte identity. Importantly, all hemocyte types, except for lamellocytes, were able to divide after wasp infection, contributing to the increased hemocyte numbers after infection. We conclude that orchestrated differentiation and division of different hemocyte types in circulation and in sessile compartment is key to a successful immune response against parasitoid wasps.

Anderl, Ines

Abstract [en]

During the last 40 years, Drosophila melanogaster has become an invaluable tool in understanding innate immunity. The innate immune system of Drosophila consists of a humoral and a cellular component. While many details are known about the humoral immune system, our knowledge about the cellular immune system is comparatively small. Blood cells or hemocytes constitute the cellular immune system. Three blood types have been described for Drosophila larvae. Plasmatocytes are phagocytes with a plethora of functions. Crystal cells mediate melanization and contribute to wound healing. Plasmatocytes and crystal cells constitute the blood cell repertoire of a healthy larva, whereas lamellocytes are induced in a demand-adapted manner after infection with parasitoid wasp eggs. They are involved in the melanotic encapsulation response against parasites and form melanotic nodules that are also referred to as tumors.

In my thesis, I focused on unraveling the mechanisms of how the immune system orchestrates the cellular immune response. In particular, I was interested in the hematopoiesis of lamellocytes.

In Article I, we were able to show that ectopic expression of key components of a number of signaling pathways in blood cells induced the development of lamellocytes, led to a proliferative response of plasmatocytes, or to a combination of lamellocyte activation and plasmatocyte proliferation.

In Article II, I combined newly developed fluorescent enhancer-reporter constructs specific for plasmatocytes and lamellocytes and developed a “dual reporter system” that was used in live microscopy of fly larvae. In addition, we established flow cytometry as a tool to count total blood cell numbers and to distinguish between different blood cell types. The “dual reporter system” enabled us to differentiate between six blood cell types and established proliferation as a central feature of the cellular immune response. The combination flow cytometry and live imaging increased our understanding of the tempo-spatial events leading to the cellular immune reaction.

In Article III, I developed a genetic modifier screen to find genes involved in the hematopoiesis of lamellocytes. I took advantage of the gain-of-function phenotype of the Tl10b mutation characterized by an activated cellular immune system, which induced the formation blood cell tumors. We screened the right arm of chromosome 3 for enhancers and suppressors of this mutation and uncovered ird1.

Finally in Article IV, we showed that the activity of the Toll signaling pathway in the fat body, the homolog of the liver, is necessary to activate the cellular immune system and induce lamellocyte hematopoiesis.